Hydrolysis is the breaking of a covalent bond in a substrate by introduction of a water molecule. The reaction involves a nucleophilic attack by the water molecule's oxygen atom on a target bond in the substrate. The water molecule is split across the target bond, breaking the bond and generating two product molecules. Hydrolytic enzymes participate in reactions essential to functions such as cell signaling, cell proliferation, inflammation, apoptosis, secretion and excretion. Hydrolytic enzymes are involved in key steps in disease processes involving these functions. Hydrolytic enzymes, or hydrolases, may be grouped by substrate specificity into classes including aminohydrolases, phospholipases, carboxyl-esterases, phosphodiesterases, lysozymes, glycosidases, glyoxalases, sulfatases, phosphohydrolases, and serine hydrolases.
NG, NG-dimethylarginine dimethylaminohydrolase (DDAH) is an enzyme that hydrolyzes the endogenous nitric oxide synthase (NOS) inhibitors. NG-monomethyl-arginine and NG, NG-dimethyl-L-arginine to L-citrulline. Inhibiting DDAH can cause increased intracellular concentration of NOS inhibitors to levels sufficient to inhibit NOS. Therefore, DDAH inhibition may provide a method of NOS inhibition and changes in the activity of DDAH could play a role in pathophysiological alterations in nitric oxide generation (MacAllister, R. J., et al. (1996) Br. J. Pharmacol. 119: 1533-1540). DDAH was found in neurons displaying cytoskeletal abnormalities and oxidative stress in Alzheimer's disease. In age-matched control cases, DDAH was not found in neurons. This suggests that oxidative stress- and nitric oxide-mediated events play a role in the pathogenesis of Alzheimer's disease (Smith, M. A., et al. (1998) Free Radic. Biol. Med. 25: 898-902).
Dipeptidyl peptidase III is an enzyme that catalyzes the release of an N-terminal dipeptide from a peptide of four or more residues. It is localized to the cytosol and is active at neutral pH. It is inactive oh Glu(4), Gly(4), and bonds involving proline. (See ExPasy—ENZYME, EC 3.4.14.4.)
Peptide deformylase hydrolyzes the formyl group at the N-terminus of newly synthesized polypeptides in prokaryotes. Deletion of the gene encoding peptide deformylase is lethal in E. coli. This lethality makes peptide deformylase a target for the design of new antibiotics (Becker, A. et al. (1998) J. Biol. Chem. 273:11413-11416 and Rajagopalan, P. T. R. and Pei, D. (1998) J. Biol. Chem. 273:22305-22310).
Trehalase is an enzyme that hydrolyzes trehalose, a protein that is thought to play a role in thermotolerance and dessication tolerance in yeast. Neutral trehalase is localized in the cytosol, while acid trehalase is localized in the vacuole. There is strong evidence that it is the neutral trehalase that hydrolyzes trehalose in intact cells. Evidence also suggests that the enhanced thermotolerance due to increased levels of trehalase is not due to the accumulation of trehalose. Trehalase may interact with heat shock protein 70 (Nwaka, S., et al. (1995) J. Biol. Chem. 270:10193-10198).
Phosphodiesterases catalyze the hydrolysis of one of the two ester bonds in a phosphodiester compound. Phosphodiesterases are, therefore, crucial to a variety of cellular processes. Phosphodiesterases include DNA and RNA endo- and exo-nucleases, which are essential to cell growth and replication as well as protein synthesis.
Pancreatic lipase and colipase form a complex that plays a key role in dietary fat digestion by converting insoluble long chain triacylgycerols into more polar molecules able to cross the brush border of intestinal cells. Colipase hinds to the C-terminal domain of lipase. In solution, this interaction involves the formation of an ion pair between a glutamic acid residue of colipase and a lysine residue of lipase. These residues are strictly conserved among species (Ayvazian, L., et. al. (1998) J. Biol. Chem. 273(50): 33604-33609). Colipase appears to overcome the inhibitory effects of bile salts on pancreatic lipase (Online Mendelian Inheritance in Man (OMIM) 246600). Diacyglycerol lipase hydrolyzes triacylglycerol, diacylglycerol and other low-density lipoproteins (ExPASy—ENZYME, EC 3.1.1.34).
Carboxylesterases are proteins that hydrolyze carboxylic esters and are classified into three categories—A, B, and C. Most type-B carboxylesterases are evolutionarily related and are considered to comprise a family of proteins. The type-B carboxylesterase family of proteins includes vertebrate acetylcholinesterase, mammalian liver microsomal carboxylesterase, mammalian bile-salt-activated lipase, and duck fatty acyl-CoA hydrolase. Some members of this protein family are not catalytically active but contain a domain related evolutionarily to other type-B carboxylesterases, such as thyroglobulin and Drosophila protein neuractin. The active site of carboxylesterases involves three residues: a serine, a glutamate or aspartate, and a histidine. The sequence surrounding this catalytic site is well conserved and can be used as a signature pattern (PROSITE: PDOC00112).
Lysozyme c superfamily consists of conventional lysozymes c, calcium-binding lysozymes c, and α-lactalbumin (Prager, E. M. and Jolles, P. (1996) EXS 75:9-31). The proteins in this superfamily have 35-40% sequence homology and share a common three dimensional fold, but can have different functions. Lysozymes bind and cleave the glycosidic bond linkage in sugars (Iyer, L. K. and Qasba, P. K. (1999) Protein Eng. 12:129-139). Lysozymes c are ubiquitous in a variety of tissues and secretions and can lyse the cell walls of ceratin bacteria (McKenzie, H. A. (1996) EXS 75: 365-409). Alpha-lactalbumin is a metallo-protein that binds calcium and participates in the synthesis of lactose (Iyer, L. K and Qasba, P. K. (1999) Protein Eng. 12: 129-139). Alpha-lactalbumin occurs in mammalian milk and colostrum (McKenzie, supra.).
The glyoxylase system consists of glyoxalase I, which catalyzes the formation of S-D-lactoylglutathione from methyglyoxal, a side product of triose-phosphate energy metabolism, and glyoxylase II, which hydrolyzes S-D-lactoylglutathione to D-lactic acid and reduced glutathione. Methyglyoxal levels are elevated during hyperglycemia, likely due to increased triose-phosphate energy metabolism. Elevated levels of glyoxylase II activity have been found in human and in a rat model of non-insulin-dependent diabetes mellitus. The glyoxylase system has been implicated in the detoxification of bacterial toxins, and in the control of cell proliferation and microtubule assembly. Elevated levels of S-D-lactoylglutathione, the substrate of glyoxylase II, induced growth arrest and toxicity in HL60 cells. Thus, the glyoxylase system, and glyoxylase II in particular, may be associated with cell proliferation and autoimmune system disorders such as diabetes.
Sulfatases are members of a highly conserved gene family that share extensive sequence homology and a high degree of structural similarity. Sulfatases catalyze the cleavage of sulfate esters. To perform this function, sulfatases undergo a unique posttranslational modification in the endoplasmic reticulum that involves the oxidation of a conserved cysteine residue. A human disorder called multiple sulfatase deficiency is due to a defect in this posttranslational modification step, leading to inactive sulfatases (Recksiek, M., et al. (1998) J. Biol. Chem. 273: 6096-6103).
Phosphohydrolases are enzymes that catalyze the hydrolysis of phosphate esters. Some phosphohydrolases contain a mutT domain signature sequence. MutT is a protein involved in the GO system responsible for removing an oxidatively damaged form of guanine from DNA. A region of about 40 amino acid residues, found in the N-terminus of mutT, is also found in other proteins, including some phosphohydrolases (PROSITE: PDOC00695).
Phosphatidic acid phosphohydrolases (PAPs) catalyze the de phophorylation of phosphatidic acid to form diacylglycerol. The hydrolysis of phosphatidic acid by PAP terminates the signaling functions of phophatidic acid and, by generating diacylglycerol, activates Ca2+- and phospholipid-dependent protein kinase C enzymes. PAP-2 is localized to the plasma membrane and is independent of Mg2+. It may play a role in modulating the signaling functions of phosphatidic acid, lysophosphatidic acid, and sphingomyelin derived lipid phosphomonoesters. Three isozymes of PAP have been found in humans to date: PAP-2a, PAP-2b, and PAP-2c. (See, Roberts, R. et al. (1998) J. Biol. Chem. 273:22059-22067.)
Glycosidases catalyze the cleavage of hemiacetyl bonds of glycosides, which are compounds that contain one or more sugar. Mammalian beta-galactosidase removes the terminal galactose from gangliosides, glycoproteins, and glycosaminoglycans. Beta-galactosidases belong to family 35 in the classification of glycosyl hydrolases. Deficiency of this enzyme is associated with the genetic disease GM1-gangliosidosis known as Morquio disease type B (PROSITE: PDOC00910).
Serine hydrolases are a functional class of hydrolytic enzymes that contain a serine residue in their active site. This class of enzymes contains proteinases, esterases, and lipases which hydrolyze a variety of substrates and, therefore, have different biological roles. Proteins in this superfamily can be further grouped into subfamilies based on substrate specificity or amino acid similarities (Puente, X. S. and Lopez-Ont, C. (1995) J. Biol. Chem. 270: 12926-12932).
The discovery of new hydrolytic enzymes and the polynucleotides encoding them satisfies a need in the art by providing new compositions which are useful in the diagnosis, prevention, and treatment of neurological disorders, immune system disorders, genetic disorders, and cell proliferation disorders, including cancer.